RESUMO
The polyunsaturated fatty acid (PUFA) synthases from deep-sea bacteria invariably contain multiple acyl carrier protein (ACP) domains in tandem. This conserved tandem arrangement has been implicated in both amplification of fatty acid production (additive effect) and in structural stabilization of the multidomain protein (synergistic effect). While the more accepted model is one in which domains act independently, recent reports suggest that ACP domains may form higher oligomers. Elucidating the three-dimensional structure of tandem arrangements may therefore give important insights into the functional relevance of these structures, and hence guide bioengineering strategies. In an effort to elucidate the three-dimensional structure of tandem repeats from deep-sea anaerobic bacteria, we have expressed and purified a fragment consisting of five tandem ACP domains from the PUFA synthase from Photobacterium profundum. Analysis of the tandem ACP fragment by analytical gel filtration chromatography showed a retention time suggestive of a multimeric protein. However, small angle X-ray scattering (SAXS) revealed that the multi-ACP fragment is an elongated monomer which does not form a globular unit. Stokes radii calculated from atomic monomeric SAXS models were comparable to those measured by analytical gel filtration chromatography, showing that in the gel filtration experiment, the molecular weight was overestimated due to the elongated protein shape. Thermal denaturation monitored by circular dichroism showed that unfolding of the tandem construct was not cooperative, and that the tandem arrangement did not stabilize the protein. Taken together, these data are consistent with an elongated beads-on-a-string arrangement of the tandem ACP domains in PUFA synthases, and speak against synergistic biocatalytic effects promoted by quaternary structuring. Thus, it is possible to envision bioengineering strategies which simply involve the artificial linking of multiple ACP domains for increasing the yield of fatty acids in bacterial cultures.
Assuntos
Proteína de Transporte de Acila/química , Ácido Graxo Sintases/química , Algoritmos , Sequência de Aminoácidos , Modelos Moleculares , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Photobacterium/enzimologia , Desnaturação Proteica , Estrutura Terciária de Proteína , Soluções , TemperaturaRESUMO
Approximately 30-50% of the >30 million HIV-infected subjects develop neurological complications ranging from mild symptoms to dementia. HIV does not infect neurons, and the molecular mechanisms behind HIV-associated neurocognitive decline are not understood. There are several hypotheses to explain the development of dementia in HIV(+) individuals, including neuroinflammation mediated by infected microglia and neuronal toxicity by HIV proteins. A key protein associated with the neurological complications of HIV, gp120, forms part of the viral envelope and can be found in the CSF of infected individuals. HIV-1-gp120 interacts with several receptors including CD4, CCR5, CXCR4, and nicotinic acetylcholine receptors (nAChRs). However, the role of nAChRs in HIV-associated neurocognitive disorder has not been investigated. We studied the effects of gp120(IIIB) on the expression and function of the nicotinic receptor α7 (α7-nAChR). Our results show that gp120, through activation of the CXCR4 chemokine receptor, induces a functional up-regulation of α7-nAChRs. Because α7-nAChRs have a high permeability to Ca(2+), we performed TUNEL staining to investigate the effects of receptor up-regulation on cell viability. Our data revealed an increase in cell death, which was blocked by the selective antagonist α-bungarotoxin. The in vitro data are supported by RT-PCR and Western blot analysis, confirming a remarkable up-regulation of the α7-nAChR in gp120-transgenic mice brains. Specifically, α7-nAChR up-regulation is observed in mouse striatum, a region severely affected in HIV(+) patients. In summary, CXCR4 activation induces up-regulation of α7-nAChR, causing cell death, suggesting that α7-nAChR is a previously unrecognized contributor to the neurotoxicity associated with HIV infection.
Assuntos
Complexo AIDS Demência/metabolismo , Corpo Estriado/metabolismo , Proteína gp120 do Envelope de HIV/metabolismo , HIV-1/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Receptores CXCR4/metabolismo , Receptores Nicotínicos/metabolismo , Complexo AIDS Demência/genética , Animais , Bungarotoxinas/farmacologia , Morte Celular/genética , Corpo Estriado/virologia , Proteína gp120 do Envelope de HIV/genética , HIV-1/genética , Humanos , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Receptores CXCR4/genética , Receptores Nicotínicos/genética , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética , Receptor Nicotínico de Acetilcolina alfa7RESUMO
The ameloblastoma according to the classification of odontogenic tumors by WHO in 2005, is classified as a benign neoplasm of odontogenic epithelial origin. One to three percent of tumors and cysts of the jaws are comprised of ameloblastomas. The tumor is locally aggressive, but often asymptomatic, showing a slow growth which is manifested as a facial swelling or radiographic incidental finding. On clinical examination, the tumor can cause symptoms such as pain, ulceration, tooth mobility, root resorption and malocclusion. Ameloblastomas have a high rate of recurrence if not completely removed. It occurs in almost all age groups, but is mainly diagnosed in the third or fourth decade of life. The tumor is very rare in children. We present an unusual case of a solid/multicystic ameloblastoma of the mandible in a 10-year-old girl. In addition, a brief review of the literature on reported cases of this pathology in children is also presented...
El ameloblastoma según la clasificación de tumores odontogénicos de la OMS del 2005 lo clasifica como una neoplasia benigna de origen epitelial odontogénico. Compromete el 1-3% de neoplasias y quistes maxilares. El tumor es agresivo localmente, pero muchas veces asintomático; presenta un lento crecimiento que se manifiesta como un aumento de volumen facial o un hallazgo incidental radiográfico. Al examen clínico el tumor puede causar síntomas como dolor, ulceración, reabsorción radicular con movilidad dentaria y maloclusión. El ameloblastoma posee gran tasa de recurrencia si no es totalmente removido. Se presenta en casi todos los grupos etarios pero principalmente se diagnostica en la tercera o cuarta década de vida, el tumor es muy poco común en niños. El tratamiento del ameloblastoma es controversial y debido a la distinta incidencia y comportamiento en niños, hace las consideraciones quirúrgicas diferentes a los adultos. Por lo que presentamos un inusual caso de un ameloblastoma solido/multiquístico mandibular en una niña de 10 años. Además de una breve revisión de la literatura sobre casos reportados de esta patología en niños...